The utilization of phosphate ions from hydroxypapatite (tooth enamel) by oral bacteria may be significant in the carious process. Single and dual in vivo labeling of rat enamel (Ca45 and P32) will provide a method for studying the fate of these ions in an in vitro culture system designed so that cariogenic bacteria can utilize enamel phosphate as substrate. Indvidual components of the culture system will be analyzed to determine the disposition of the calcium and phosphate ions during bacterial metabolism. An isolated from deep dentinal caries (L. casei) that forms large intracellular granules will be used as a prototype for these experiments. In previous experiments, using the prototype organism, it was noted that the granules form during the late log or early stationary phase of growth and unbalanced growth favors their formation. Intact granules were isolated from culture grown samples of this organism using chromatography and analysis indicated a phosphate content above 70 percent. The granules were volatilized by a high density electron beam in the electronmicroscope and are probably polyphosphate in nature. In proposed studies, the optimum growth conditions for the formation of these granules will be determined using a relatively defined medium and varying nutritional and physical parameters. After these optimums are established Ca45 and/or P32 labeled hydroxyapatite will be substituted for phosphate in the medium. When growth is completed, the supernatant fluid, the individual cells and isolated granules will be analyzed for labeled ions. In addition, the chemistry and structure of the granules will be determined, the transferring enzymes identified and the effect of inhibitors examined. The methods of procedure developed with the prototype organism will be used to study strains of S. mutans and S. salivarius described as carciogenic by other laboratories.